Department of Plant Medicals, Andong National University, Andong, 36729, South Korea.
Industry Academy Cooperation Foundation, Andong National University, Andong, 36729, South Korea.
Dev Comp Immunol. 2020 Jun;107:103643. doi: 10.1016/j.dci.2020.103643. Epub 2020 Feb 14.
Eicosanoids mediate both cellular and humoral immune responses in insects. Epoxyeicosatrienoic acids (EETs) are a group of eicosanoids containing epoxide formed by epoxygenase (EPX) activity of cytochrome P450 (CYP). Although EETs have been considered to mediate immune responses in some insects, their synthetic machinery was little understood in insects. This study monitored EETs in a lepidopteran insect, Spodoptera exigua, immunized with bacteria and found all four EETs (5,6-EET, 8,9-EET, 11,12-EET, and 14,15-EET) from larval fat body at 247-1,736 pg/g levels. Then to predict EPXs, 140 CYPs were collected from S. exigua transcriptomes and compared with human EPXs. Four CYPs (SeEPX1-SeEPX4) sharing homologies with human EPXs were chosen and assessed in subsequent expression and functional analyses. All four EPXs were expressed in all development stages. In larval stage, all four EPXs were expressed in immune-associated tissues such as fat body and hemocytes. Furthermore, their expression levels were highly enhanced by bacterial challenge in different tissues. RNA interference (RNAi) using gene-specific double stranded RNA injection suppressed their expression levels by more than 55%. RNAi treatments interfered with hemocyte-spreading behavior and nodule formation upon bacterial challenge except RNAi treatment against SeEPX2. All four EETs stimulated cellular immune response measured by nodule formation in S. exigua. The suppressed immune responses by the RNAi treatments against three SeEPXs were rescued by the addition of 8,9-EET. However, other three EETs gave their specific rescue effect depending on SeEPX types under RNAi. In humoral immune response, all four RNAi treatments suppressed expression of antimicrobial peptide genes. This study reports the presence of all four EETs in larval fat body of S. exigua and suggests that four SeEPXs are associated with immune responses mediated by EETs.
类二十烷酸在昆虫中既介导细胞免疫反应，也介导体液免疫反应。环氧化物二十碳三烯酸（EETs）是一组包含环氧化物的类二十烷酸，由细胞色素 P450（CYP）的环氧合酶（EPX）活性形成。尽管 EETs 已被认为在某些昆虫中介导免疫反应，但它们的合成机制在昆虫中知之甚少。本研究监测了经细菌免疫的鳞翅目昆虫 S. exigua 幼虫脂肪体中的 EETs，发现所有四种 EETs（5,6-EET、8,9-EET、11,12-EET 和 14,15-EET）的含量在 247-1,736 pg/g 水平。然后，为了预测 EPX，从 S. exigua 转录组中收集了 140 个 CYP，并与人类 EPX 进行了比较。选择了与人类 EPX 具有同源性的四个 CYP（SeEPX1-SeEPX4），并在随后的表达和功能分析中进行了评估。这四个 EPX 在所有发育阶段都有表达。在幼虫阶段，这四个 EPX 在免疫相关组织如脂肪体和血细胞中都有表达。此外，它们的表达水平在不同组织中受到细菌挑战的强烈增强。使用基因特异性双链 RNA 注射的 RNA 干扰（RNAi）将其表达水平抑制超过 55%。RNAi 处理干扰了血细胞扩散行为和细菌挑战后的结节形成，除了针对 SeEPX2 的 RNAi 处理。所有四种 EET 都刺激了 S. exigua 中结节形成所测量的细胞免疫反应。针对三个 SeEPX 的 RNAi 处理对免疫反应的抑制作用可通过添加 8,9-EET 得到挽救。然而，在 RNAi 下，其他三种 EET 根据 SeEPX 类型发挥了特定的挽救作用。在体液免疫反应中，所有四种 RNAi 处理都抑制了抗菌肽基因的表达。本研究报告了 S. exigua 幼虫脂肪体中所有四种 EET 的存在，并表明四个 SeEPX 与 EET 介导的免疫反应有关。
